Estimation of total rate of formation of nitric oxide in the rat.
Nitric oxide (NO) is a powerful mediator with important actions in several organ systems. NO is synthesized during the enzymatic conversion of l-arginine and molecular oxygen to L-citrulline. About 90% of the NO formed is degraded to nitrate. Utilizing this information we have developed a method for assessment of the total rate of formation of NO in the rat. Male Wistar rats were kept in a closed-cage system allowing controlled breathing of a mixture of 18O2 and 16O2 in N2 for up to 5h. Blood samples for mass spectrometric analysis of nitrate residues with varying numbers of 18O atoms incorporated were drawn before and during the exposure to 18O2. By comparing the relative incorporation of 18O into nitrate residues to the 16O2/18O2 ratio in the breathing gas mixture in the cage system it was possible to calculate the absolute rate of NO formation in the animal. The rate of formation of NO in anaesthetized rats ranged from 0.33 to 0.85 micromol.kg-1.h-1. The rate of formation did not differ significantly in rats which were awake during the experiment (range 0.36-0.72 micromol.kg-1.h-1). The L-arginine analogue Nomega-nitro-L-arginine methyl ester (L-NAME) dose-dependently inhibited the formation of NO, at a dose of 100mg/kg by more than 99%. The technique presented allows estimation of the total rate of formation of NO in vivo in rats. Application of the technique may yield important information about the physiological and pathophysiological roles of NO. It may also be utilized to evaluate the effect of pharmacological treatment on NO formation.
Sakinis
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Department of Clinical Physiology, Göteborg University, Sahlgrenska University Hospital, S-413 45 Göteborg, Sweden.
Wennmalm
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D000818:Animals; D004791:Enzyme Inhibitors / Q000494:pharmacology; D008297:Male; D008722:Methods; D019331:NG-Nitroarginine Methyl Ester / Q000494:pharmacology; D009566:Nitrates / Q000378:metabolism; D009569:Nitric Oxide / Q000378:metabolism*; D019001:Nitric Oxide Synthase / Q000037:antagonists & inhibitors; D010100:Oxygen / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D014851:Wakefulness
D004791:Enzyme Inhibitors; D009566:Nitrates; D009569:Nitric Oxide; D019001:Nitric Oxide Synthase; D010100:Oxygen; D019331:NG-Nitroarginine Methyl Ester